Pathophysiology of visceral leishmaniasis (VL) is not fully understood and it has been widely accepted that the parasitic components and host immune response both contribute to the perpetuation of the disease. Host alterations during leishmaniasis is a feebly touched area that needs to be explored more to better understand the VL prognosis and diagnosis, which are vital to reduce mortality and post-infection sequelae. To address this, we performed untargeted metabolomics of Leishmania donovani (Ld) infected, uninfected and treated BALB/c mice's tissues and biofluids to elucidate the host metabolome changes using gas chromatography-mass spectrometry. Univariate and multivariate data treatments provided numerous significant differential hits in several tissues like the brain, liver, spleen and bone marrow. Differential modulations were also observed in serum, urine and fecal samples of Ld-infected mice, which could be further targeted for biomarker and diagnostic validations. Several metabolic pathways were found to be upregulated/downregulated in infected (TCA, glycolysis, fatty acids, purine and pyrimidine, etcetera) and treated (arginine, fumaric acid, orotic acid, choline succinate, etcetera) samples. Results also illustrated several metabolites with different pattern of modulations in control, infected and treated samples as well as in different tissues/biofluids; for e.g. glutamic acid identified in the serum samples of infected mice. Identified metabolites include a range of amino acids, saccharides, energy-related molecules, etcetera. Furthermore, potential biomarkers have been identified in various tissues-arginine and fumaric acid in brain, choline in liver, 9-(10) EpOME in spleen and bone marrow, N-acetyl putrescine in bone marrow, etcetera. Among biofluids, glutamic acid in serum, hydrazine and deoxyribose in urine and 3-Methyl-2-oxo pentanoic acid in feces are some of the potential biomarkers identified. These metabolites could be further looked into for their role in disease complexity or as a prognostic marker. The presented profiling approach allowed us to attain a metabolic portrait of the individual tissue/biofluid modulations during VL in the host and represent a valuable system readout for further studies. Our outcomes provide an improved understanding of perturbations of the host metabolome interface during VL, including identification of many possible potential diagnostic and therapeutic targets.

Mycoplasma pneumoniae (Mp) is a highly contagious respiratory pathogen responsible for human community-acquired pneumonia. The number of antibiotic-resistant Mp strains is increasing; therefore, to develop novel therapeutics, it is crucial to precisely understand the pathogenesis of mycoplasma pneumonia. Herein, we examined the susceptibility and response to Mp among eight inbred mouse strains. Following infection, the bacterial load in the bronchoalveolar lavage fluid (BALF) from DBA/2 mice was higher than that in the other tested strains such as BALB/c mice, which are frequently used in Mp research. In contrast, the numbers of CD45+ immune cells and neutrophils in BALF were comparable between BALB/c and DBA/2 mice, with lower numbers observed in C57BL/6J and CBA/N mice than in BALB/c mice. Among the tested strains, the BALF level of interleukin 12 subunit p40 was highest in DBA/2 mice; however, significant differences in other cytokines levels were not observed between BALB/c and DBA/2 mice. After Mp infection, Mp-specific Th1 and Th17 responses were significantly enhanced in DBA/2 mice when compared with BALB/c mice. Furthermore, prior infection with Mp increased the number of neutrophils in BALF after the reinfection of DBA/2 mice through an Mp-specific CD4+ T cell-dependent mechanism. Thus, DBA/2 may be an appropriate strain for evaluating Mp infection. Moreover, a comparison of responses revealed by various inbred mouse strains could be useful for elucidating the pathogenesis of Mycoplasma pneumonia.

The surge of multidrug-resistant fungal pathogens, especially Candida auris, poses significant threats to global public health. Candida auris exhibits resistance to multiple antifungal drugs, leading to major outbreaks and a high mortality rate. With an urgent call for innovative therapeutic strategies, this study focused on the regulation and pathobiological significance of secreted aspartyl proteinases (SAPs) in C. auris, as these enzymes play pivotal roles in the virulence of some fungal species. We delved into the Ras/cAMP/PKA signaling pathway's influence on SAP activity in C. auris. Our findings underscored that the Ras/cAMP/PKA pathway significantly modulates SAP activity, with PKA catalytic subunits, Tpk1 and Tpk2, playing a key role. We identified a divergence in the SAPs of C. auris compared to Candida albicans, emphasizing the variation between Candida species. Among seven identified secreted aspartyl proteases in C. auris (Sapa1 to Sapa7), Sapa3 emerged as the primary SAP in the pathogen. Deletion of Sapa3 led to a significant decline in SAP activity. Furthermore, we have established the involvement of Sapa3 in the biofilm formation of C. auris. Notably, Sapa3 was primarily regulated by Tpk1 and Tpk2. Deletion of SAPA3 significantly reduced C. auris virulence, underscoring its pivotal role in C. auris pathogenicity. The outcomes of this study provide valuable insights into potential therapeutic targets, laying the groundwork for future interventions against C. auris infection.

Every year, up to 90,000 new cases of Visceral Leishmaniasis and 30,000 resultant deaths are estimated to occur worldwide. Such numbers give relevance to the continuous study of this complex form of the disease: a zoonosis and an anthroponosis; two known etiological agents (Leishmania infantum and L. donovani, respectively); with an estimated average ratio of 1 symptomatic per 10 asymptomatic individuals; and sometimes associated with atypical clinical presentations. This complexity, which results from a long co-evolutionary process involving vector-host, host-pathogen, and pathogen-vector interactions, is still not completely understood. The determinants of visceralization are not fully defined and the dichotomy resistance vs. susceptibility remains unsolved, translating into obstacles that delay the progress of global disease control. Inbred mouse models, with different susceptibility patterns to Leishmania infection, have been very useful in exploring this dichotomy. BALB/c and C57BL/6 mice were described as susceptible strains to L. donovani visceral infection, while SV/129 was considered resistant. Here, we used these three mouse models, but in the context of L. infantum infection, the other Leishmania species that cause visceral disease in humans, and dynamically compared their local and systemic infection-induced immune responses in order to establish a parallel and to ultimately better understand susceptibility vs. resistance in visceral leishmaniasis. Overall, our results suggest that C57BL/6 mice develop an intermediate "infection-phenotype" in comparison to BALB/c and SV/129 mouse strains, considering both the splenic parasite burden and the determined target organs weights. However, the immune mechanisms associated with the control of infection seem to be different in each mouse strain. We observed that both BALB/c and SV/129, but not C57BL/6 mice, show an infection-induced increase of splenic T follicular helper cells. On the other hand, differences detected in terms of CD21 expression by B cells early after infection, together with the quantified anti-Leishmania specific antibodies, suggest that SV/129 are faster than BALB/c and C57BL/6 mice in the assembly of an efficient B-cell response. Additionally, we observed an infection-induced increase in polyfunctional CD4+ T cells in the resistant SV/129 model, opposing an infection-induced increase in CD4+IL-10+ cells in susceptible BALB/c mice. Our data aligns with the observations reported for L. donovani infection and suggest that not only a single mechanism, but an interaction of several could be necessary for the control of this parasitic disease.

Chagas disease (ChD) is a chronic infection caused by Trypanosoma cruzi. This highly diverse intracellular parasite is classified into seven genotypes or discrete typing units (DTUs) and they overlap in geographic ranges, vectors, and clinical characteristics. Although studies have suggested that ChD progression is due to a decline in the immune response quality, a direct relationship between T cell responses and disease outcome is still unclear. To investigate the relationship between parasite control and immune T cell responses, we used two distinct infection approaches in an animal model to explore the histological and parasitological outcomes and dissect the T cell responses in T. cruzi-infected mice. First, we performed single infection experiments with DA (TcI) or Y (TcII) T. cruzi strains to compare the infection outcomes and evaluate its relationship with the T cell response. Second, because infections with diverse T. cruzi genotypes can occur in naturally infected individuals, mice were infected with the Y or DA strain and subsequently reinfected with the Y strain. We found different infection outcomes in the two infection approaches used. The single chronic infection showed differences in the inflammatory infiltrate level, while mixed chronic infection by different T. cruzi DTUs showed dissimilarities in the parasite loads. Chronically infected mice with a low inflammatory infiltrate (DA-infected mice) or low parasitemia and parasitism (Y/Y-infected mice) showed increases in early-differentiated CD8+ T cells, a multifunctional T cell response and lower expression of inhibitory receptors on CD8+ T cells. In contrast, infected mice with a high inflammatory infiltrate (Y-infected mice) or high parasitemia and parasitism (DA/Y-infected mice) showed a CD8+ T cell response distinguished by an increase in late-differentiated cells, a monofunctional response, and enhanced expression of inhibitory receptors. Overall, our results demonstrated that the infection outcomes caused by single or mixed T. cruzi infection with different genotypes induce a differential immune CD8+ T cell response quality. These findings suggest that the CD8+ T cell response might dictate differences in the infection outcomes at the chronic T. cruzi stage. This study shows that the T cell response quality is related to parasite control during chronic T. cruzi infection.

Hantaan virus (HTNV), the causative pathogen of hemorrhagic fever with renal syndrome (HFRS), is a negative RNA virus belonging to the Orthohantaviridae family. HTNV envelope glycoprotein (GP), encoded by the genomic medium segment, is immunogenic and is therefore a promising vaccine candidate. Major histocompatibility complex class I (MHC-I) epitopes derived from HTNV has been extensively studied, but little is known of MHC-II epitopes. In silico predictions based on four databases indicated that the full-length HTNV GP has 1121 15-mer epitopes, of which 289 had a high score for binding to the human and murine MHC-II superfamily. It found that epitope ILTVLKFIANIFHTS could potentially bind most MHC-II molecules covering human and murine haplotypes. Dominant epitopes were validated by enzyme-linked immunospot assay of splenocytes from immunized mice; 6 of 10 epitopes supported the predictions including TATYSIVGPANAKVP, TKTLVIGQCIYTITS, FSLLPGVAHSIAVEL, CETYKELKAHGVSCP, CGLYLDRLKPVGSAY, and NLGENPCKIGLQTSS. Conservation analysis of dominant epitopes revealed host-virus interactions without geographic stratification, thus meeting the requirements of candidate vaccines for large-population prophylaxis. These findings provide insight into hantavirus antigenicity and suggest that vaccines targeting MHC-II could provide immune protection in large population to complement symptomatic therapies for the treatment of HFRS.

Zinc oxide nanoparticles (ZnO-NPs) have garnered considerable interest in biomedical research primarily owing to their prospective therapeutic implications in combatting pathogenic diseases and microbial infections. The primary objective of this study was to examine the biosynthesis of zinc oxide nanowhiskers (ZnO-NWs) using chicken egg white (albumin) as a bio-template. Furthermore, this study aimed to explore the potential biomedical applications of ZnO NWs in the context of infectious diseases.

Infection with the apicomplexan protozoan parasite Toxoplasma gondii is an ongoing public health problem. The parasite's ability to invade and replicate within the host cell is dependent on many effectors, such as dense granule proteins (GRAs) released from the specialized organelle dense granules, into host cells. GRAs have emerged as important determinants of T. gondii pathogenesis. However, the functions of some GRAs remain undefined. In this study, we used CRISPR-Cas9 technique to disrupt 17 GRA genes (GRA11, GRA12 bis, GRA13, GRA14, GRA20, GRA21, GRA28-31, GRA33-38, and GRA40) in the virulent T. gondii RH strain. The CRISPR-Cas9 constructs abolished the expression of the 17 GRA genes. Functional characterization of single ΔGRA mutants was achieved in vitro using cell-based plaque assay and egress assay, and in vivo in BALB/c mice. Targeted deletion of these 17 GRA genes had no significant effect neither on the in vitro growth and egress of the mutant strains from the host cells nor on the parasite virulence in the mouse model of infection. Comparative analysis of the transcriptomics data of the 17 GRA genes suggest that GRAs may serve different functions in different genotypes and life cycle stages of the parasite. In sum, although these 17 GRAs might not be essential for RH strain growth in vitro or virulence in mice, they may have roles in other strains or parasite stages, which warrants further investigations.

In human cutaneous leishmaniasis (HCL) caused by Leishmania (L.) major, the cutaneous lesions heal spontaneously and induce a Th1-type immunity that confers solid protection against reinfection. The same holds true for the experimental leishmaniasis induced by L. major in C57BL/6 mice where residual parasites persist after spontaneous clinical cure and induce sustainable memory immune responses and resistance to reinfection. Whether residual parasites also persist in scars of cured HCL caused by L. major is still unknown. Cutaneous scars from 53 volunteers with healed HCL caused by L. major were biopsied and the tissue sample homogenates were analyzed for residual parasites by four methods: i) microscope detection of amastigotes, ii) parasite culture by inoculation on biphasic medium, iii) inoculation of tissue exctracts to the footpad of BALB/c mice, an inbred strain highly susceptible to L. major, and iv) amplification of parasite kDNA by a highly sensitive real-time PCR (RT-PCR). Our results show that the scars of healed lesions of HCL caused by L. major do not contain detectable residual parasites, suggesting that this form likely induces a sterile cure at least within the scars. This feature contrasts with other Leishmania species causing chronic, diffuse, or recidivating forms of leishmaniasis where parasites do persist in healed lesions. The possibility that alternative mechanisms to parasite persistence are needed to boost and maintain long-term immunity to L. major, should be taken into consideration in vaccine development against L. major infection.